Many vertebrates have robust capacities for tissue regeneration as embryos and juveniles but this power fades during development and aging. The mechanisms of age-related decline in regenerative capacity are unknown and an area in which new approaches and model systems are necessary. This proposal develops the tunicate Ciona intestinalis as a chordate model for studying the effects of aging on tissue regeneration. Although not previously used as an aging model, Ciona has many favorable attributes for this purpose: it has powerful capacities for regeneration, which decline during aging, a sequenced genome, a collection of cDNA and ESTs covering 80% of the total genes, stable transgenic lines with tagged gene expression in specific tissues, and a short life history in which age is directly related to size. As a member of a chordate invertebrate group that is inferred to be the sister taxon of vertebrates, information obtained about the relationship between aging and regeneration in tunicates may be relevant to humans. The five specific aims of this proposal are designed to obtain a molecular and cellular understanding of defective oral siphon regeneration during aging. We will focus on the sensory pigment organs of the oral siphon, possible photoreceptors that regenerate with high fidelity in young and middle age animals, but show specific defects in old animals. The first three aims will focus on the cellular events involved in producing age-related defects in pigment organ regeneration. The first aim will investigate the effects of aging on maintenance of the stem/progenitor cell niche for pigment organ precursors and precursor cell migration to the wound site during regeneration. The second aim will focus on age-related changes in cell death and proliferation in the siphon wound site, the stem/precursor niche, and the reforming pigment organs during regeneration. The third aim explores the role of siphon nerves and the CNS in regenerative aging using ablation techniques and a transgenic line with GFP staining throughout the nervous system. The fourth and fifth aims investigate the molecular mechanisms responsible for age-related defects in pigment organ regeneration. The fourth aim will identify and determine the expression patterns of genes involved in regeneration, including components of the Notch signaling system, which preliminary studies have implicated in the regenerative process. The fifth aim will use RNAi gene silencing to establish how functional changes in regeneration genes are involved in the cellular aging processes revealed in the first three aims. This research is expected to fill a major gap in understanding how regenerative capacity declines with aging in a model chordate representing the closest living relative of vertebrates. PUBLIC HEALTH RELEVANCE: The ability to replace injured tissues fades with aging in most vertebrates, including humans. This study will develop the tunicate Ciona intestinalis, a vertebrate relative with powerful regeneration abilities that also recede with age, as a model to study the relationship between aging and tissue regeneration.